Control of Blood Glucose

Glucocorticoids act in concert with other hormones to maintain or raise blood glucose levels by (1) stimulating hepatic gluconeogenesis, (2) mobilizing gluconeogenic substrates from peripheral tissues, (3) permissively enhancing and prolonging the effects of glucagon and epinephrine on gluconeogenesis and glycogenolysis, (4) inhibiting peripheral glucose utilization, and (5) promoting liver glycogen synthesis to store substrate in preparation for acute responses to glycogenolytic agents such as glucagon and epinephrine.⁴⁵

From an evolutionary standpoint, glucocorticoids in this way support stress responses that require glucose for rapid and intense exertion, such as an encounter of prey with predator.²⁶ From a physiologic and clinical standpoint, glucocorticoids are counterregulatory hormones that protect the body from insulin-induced hypoglycemia. Both of these roles, in which glucocorticoid effects develop over the course of hours, are shared with the rapidly acting glucagon and epinephrine and to some extent with growth hormone.⁸⁴

Glucocorticoid actions interact with those of insulin during feeding and fasting in complex ways that not only maintain blood glucose but influence appetite, feeding patterns, disposal of foodstuffs, and body composition.^26,85 Antagonism with insulin in both glucose synthesis and utilization accounts at least partly for the diabetogenic actions of excessive glucocorticoids.^45,86 Studies with transgenic mice show that expression of hepatic peroxisome-proliferator–activated receptor-α (PPAR-α) may be one mechanism underlying glucocorticoid-induced hypertension and insulin resistance.⁸⁷

Gluconeogenesis

Hepatic gluconeogenesis is stimulated by glucocorticoids, mainly through the increased activities of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase. These enzymes catalyze the conversion of oxaloacetate to phosphoenolpyruvate and of glucose-6-phosphate to glucose—both rate-limiting steps in gluconeogenesis.^45,88 Glucocorticoids also regulate expression of 6-phosphofructo-2-kinase/fructose 2,6-biphosphatase, a bifunctional enzyme that controls the level of fructose-2,6-biphosphate. Fructose-2,6-biphosphate is an allosteric regulator of gluconeogenic and glycolytic enzymes. PEPCK and 6-phosphofructo-2-kinase/fructose 2,6-biphosphatase activities are controlled principally through synthesis of the enzymes.⁸⁸ On starvation, 11β-HSD1 knockout mice have diminished activation of PEPCK and glucose-6-phosphatase.⁸⁹

Control of PEPCK gene expression reflects the complexity of regulation of gluconeogenesis in the body, involving glucocorticoids, insulin, glucagon, catecholamines, cyclic adenosine monophosphate (cAMP), and retinoic acid.^88,90 In particular, glucocorticoids and insulin, by respectively promoting and indirectly disrupting association of CBP (CREB-binding protein) and RNA polymerase II with the PEPCK promoter, reciprocally regulate PEPCK gene expression. The PEPCK gene has a glucocorticoid response unit (GRU) that spans 110 base pairs. There are two GR-binding sites and four accessory factor elements, all of which are required for glucocorticoid regulation, and within the GRU are insulin-responsive and retinoic acid–responsive sequences.^88,91 The 6-phosphofructo-2-kinase/fructose 2,6-biphosphatase gene has a complex glucocorticoid response element that resembles the GRU of the PEPCK gene. Hepatocyte nuclear factor-6 (HNF-6) inhibits glucocorticoid activation of both these genes by binding to DNA and GRs. As would be expected, treatment with glucocorticoids of transgenic mice with dimerization-deficient GRs (i.e., GRs that cannot bind to GREs) failed to induce PEPCK.³⁵

Substrates for gluconeogenesis are generated by glucocorticoids through release of amino acids from muscle and other peripheral tissues and release of glycerol along with lipolysis.

Permissive actions of glucocorticoids on gluconeogenesis by glucagon and epinephrine, possibly due to enhanced responsiveness to cAMP or other intracellular mediators, are evidenced by the impairment of gluconeogenesis caused by adrenalectomy and its normalization by glucocorticoids.

Glucose Utilization

Glucocorticoid inhibition of peripheral glucose utilization can be demonstrated both in intact organisms and with isolated cells.⁹² It probably accounts for significant insulin antagonism and for the early rise in blood glucose seen after glucocorticoid treatment, and it may play a role in the release of gluconeogenic substrates from peripheral tissues. Glucose uptake is inhibited by direct glucocorticoid actions on normal skin, fibroblast, adipose tissue, adipocytes, lymphoid cells, and polymorphonuclear leukocytes. This inhibition, which requires RNA and protein synthesis, has been postulated to be mediated by a glucocorticoid-induced protein. It results mainly from translocation of glucose transporters from the plasma membrane to intracellular sites.^93,94 Glucose uptake by muscle is inhibited in intact organisms treated with glucocorticoids. This action may be indirect.

Glycogen Synthesis and Breakdown

Stimulation of glycogen synthesis by glucocorticoids takes place in the fetus and in the adult. It depends on increased synthesis of hepatic glycogen synthase and activation by dephosphorylation of its inactive form, as well as on inactivation by dephosphorylation of phosphorylase a. Some of these changes can be accounted for by increases in glycogen-bound phosphatase activity.

Stimulation of liver glycogen synthesis by stress-induced glucocorticoids can be interpreted as preparation for a subsequent challenge in which glycogen will be used for rapid conversion to glucose by glycogenolysis.²⁶ Furthermore, glucocorticoids at basal levels are required to permissively maintain epinephrine-induced glycogenolysis, which is impaired by adrenalectomy.

Fat Metabolism

Glucocorticoids, in opposition to insulin, inhibit glucose transport by adipose cells and stimulate free fatty acid release, which in humans results in an increase in plasma free fatty acids within 1 to 2 hours of administration of hormone.⁹⁵ Increased release of fatty acids also occurs after incubation of adipose tissue with glucocorticoids, an effect that is due in part to decreased reesterification resulting from the decrease in glucose uptake, and in part to increased lipolysis. Stimulation of lipolysis is largely a permissive effect seen in the presence of growth hormone and other lipolytic agents.⁹⁵ Glucocorticoids also inhibit the action of leptin and are permissive for the obesity syndrome in mutant rodents, which is ameliorated by adrenalectomy. A curious observation is that mice expressing a GR antisense construct that lowers their GR levels have as their most striking abnormality an increase in fat deposition, which can double their weight compared with normal mice. Because these mice eat less than normal, they are presumed to have increased energy efficiency.⁹⁶

Chronic stress, which increases low diurnal concentrations of glucocorticoids, has been linked to central obesity and the metabolic syndrome, a disorder that combines diabetes, insulin resistance, dyslipidemia, and hypertension.⁶¹ 11β-HSD1 knockout mice have weakened glucocorticoid-induced responses and resist hyperglycemia provoked by obesity or stress.⁸⁹ The critical importance of local glucocorticoid levels has been demonstrated with transgenic mice that overexpress 11β-HSD1 in fat cells: those mice develop central obesity and the metabolic syndrome.^45,47

Catabolic Effects

Chronic high levels of glucocorticoids lead to massive catabolic effects on proteins and other components of peripheral tissues, causing muscle wasting⁹⁷ and lipolysis with redistribution of fat. These pathologic changes are probably magnified expressions of physiologic mechanisms for generating gluconeogenic substrates and may result from interactions with insulin and other hormones.⁴⁵

Antiinflammatory and Immunosuppressive Actions

Among the major clinical applications of hormones is the use of glucocorticoids for suppression of inflammatory and immune reactions and for treatment of patients with cancers of the lymphoid system. (Evolution anticipated such a use with vaccinia virus, which encodes an enzyme, 3β-hydroxysteroid dehydrogenase, which in infected organisms enhances glucocorticoid production, suppressing the inflammatory response and increasing virulence.¹⁰³)

As already described, for years glucocorticoid suppression of inflammatory and immune reactions was believed to result from pharmacologic actions with no physiologic significance. Strong evidence, however, points to their physiologic nature. They are elicited through the same receptor-mediated genomic mechanisms as physiologic effects. Adrenalectomy or administration of the glucocorticoid antagonist mifepristone (RU486) enhance responses to inflammatory agents, showing that endogenous glucocorticoids normally control inflammation and similarly control autoimmune reactions. A striking example is the high susceptibility to arthritis of Lewis rats compared to the largely histocompatible Fischer rats after challenge with streptococcal cell wall polysaccharide (SCW).¹⁰⁴ This difference is due to a defect in biosynthesis of CRH that limits the glucocorticoid response of Lewis rats to a challenge. Lewis rats can be protected from SCW with dexamethasone, whereas Fischer rats become susceptible to SCW when pretreated with RU486. Defective HPA function may have an etiologic role in rheumatoid arthritis.¹⁰⁵ In transgenic mice with GRs that cannot dimerize and therefore cannot transactivate genes through binding to palindromic GREs, most anti-inflammatory and immunosuppressive actions of glucocorticoids remain intact, indicating that they are mediated through binding of GRs to factors such as AP-1 and NF-κB.³⁴

Glucocorticoids also exert permissive actions on the immune system, as will be described below. Stimulation of the HPA axis may mobilize immunoregulatory agents other than glucocorticoids. One already mentioned is CRH, which peripherally has proinflammatory activity.¹⁰⁴ Whether such agents normally participate in immune responses to stress is uncertain.

Effects on Leukocytes

Glucocorticoids influence most cells responsible for immune and inflammatory reactions, including lymphocytes, natural killer (NK) cells, monocytes and macrophages, dendritic cells, eosinophils, neutrophils, mast cells, and basophils. Accumulation of most of these cells is decreased at inflammatory sites, an effect that can be induced by local application of hormones, but they mobilize neutrophils from bone marrow to blood.^100,106 Glucocorticoids may have both beneficial and detrimental effects on wound healing.¹⁰⁷ Blood counts of lymphocytes, monocytes, eosinophils, and basophils drop within 1 to 3 hours of glucocorticoid administration, generally recovering in 12 to 48 hours. NK cells are unaffected, and neutrophil counts rise. CD4 or helper T cells are more sensitive to lymphopenia than are B cells, and CD8 or cytotoxic T cells are relatively insensitive. Increased neutrophil number is thought to reflect increased release of marginated cells to the circulation and increased half-life. These alterations in cell traffic probably depend on inhibiting expression of surface molecules such as endothelial leukocyte adhesion molecule (ELAM)-1 and intercellular adhesion molecule (ICAM)-1,^104,108 thus decreasing adhesion of leukocytes to endothelial and other cells.

Glucocorticoid administration usually reduces antigen- or lectin-induced mitogenesis measured with peripheral lymphocytes, an effect also observed with lymphocytes in culture. T cells are more sensitive than B cells, and helper more sensitive than cytotoxic T cells. Glucocorticoids also directly inhibit T and B cell proliferation, early B cell differentiation, NK activity, and the differentiation and function of macrophages. They inhibit antigen presentation by monocytes and by dendritic cells (the most potent antigen-presenting cells) and shift responses from T helper 1 (Th1) cells to Th2 cells¹⁰⁹ Although glucocorticoids have stimulatory effects on immunoglobulin synthesis in cell culture, in whole organisms glucocorticoids usually inhibit B cell function.

Permissive glucocorticoid actions on T cell function have been observed in human volunteers treated with lipopolysaccharide (LPS), a mediator of septic shock: when administered within 6 hours of LPS, cortisol hemisuccinate suppressed the LPS-induced increase in TNF, but when given 12 to 144 hours before LPS, it magnified the TNF response.¹¹⁰ Both in rats and in cultured splenic lymphocytes, glucocorticoids at low concentrations, presumably acting through MRs, can enhance T cell responses to concanavalin A, whereas at higher concentrations, through GRs, they suppress.¹¹¹ Hormone concentration and timing appear to be important for these actions to be displayed separately from the usually predominant suppressive effects.¹¹² Delayed-type hypersensitivity (DTH) reactions to cutaneous antigen exposure are enhanced by acute physiologic increases in glucocorticoid levels (and eliminated by adrenalectomy), but they are suppressed by chronic exposure to glucocorticoids.¹¹³ In treatment for contact hypersensitivity, the therapeutic action of glucocorticoids is exerted through macrophages and neutrophils and requires GR dimerization.¹¹⁴ The value of glucocorticoids in treatment for septic shock is controversial¹⁰⁸ (see Chapter 115).

Important effects of glucocorticoids are exerted through the innate immune system. This ancient defense system uses about 10 invariant germline-encoded toll-like receptors (TLRs) on monocytes, macrophages, and dendritic and other cells to respond to infectious molecules of microbial origin by inducing antimicrobial genes along with inflammatory cytokines and chemokines, thus stimulating leukocyte migration and triggering adaptive immune responses.¹¹⁵ LPS, the best-known stimulator of the innate system, provokes a rapid increase in glucocorticoid levels, which, in turn, protects the organism from potentially lethal effects of LPS.^108,116 The HPA response and other effects of LPS are mediated in part by the proinflammatory cytokines TNF-α, IL-1, and IL-6. In the airway epithelium, glucocorticoids may enhance innate immunity.^117,118 Glucocorticoid treatment of myeloid progenitors also enhances TLR signaling in macrophages to which they differentiate.¹¹⁹ In other circumstances, glucocorticoids inhibit TLR signaling.¹²⁰

Glucocorticoids protect against overactivity of immune reactions through several mechanisms, including suppression of production or potentially toxic activity of proinflammatory cytokines, histamine, adhesion molecules, inducible cyclooxygenase, and inducible nitric oxide synthase. Glucocorticoids also suppress expression and release of the LPS receptor CD-14. Overexpression of GRs increases resistance to LPS, thereby reducing production of IL-6.¹²¹

The anti-inflammatory cytokine IL-10 may play an important role in controlling LPS effects. Neutralization of IL-10 enhances the lethality of LPS in mice, whereas IL-10 administration reduces lethality.¹²² Optimal IL-10 production during cardiac surgery requires a surge in glucocorticoid levels.¹²³ In experimental human endotoxemia, even high doses of glucocorticoids enhance IL-10 production.¹²⁴

During development, LPS may affect the HPA axis and have long-lasting effects on immune regulation. Exposure of neonatal rats to LPS raises their adult corticosterone levels and protects the adults from adjuvant-induced arthritis.¹²⁵

Apoptosis

Glucocorticoid-induced apoptosis of thymocytes and other lymphocytes is among the most striking effects of these hormones,^12,126,127 and the underlying mechanisms of this effect are being revealed gradually.³¹ Apoptosis is directly linked to control of T cell pools in vivo, as is demonstrated with transgenic mice. Mice with increased GR expression have increased sensitivity to glucocorticoid-induced apoptosis,¹²¹ and mice with higher or lower GR levels targeted to lymphocytes have, respectively, smaller or larger T cell pools.¹²⁸ Glucocorticoid-induced apoptosis has been demonstrated with most hematologic cells, as well as with other cells such as epithelial and carcinoma cells and osteoblasts.^129,130 In some circumstances, glucocorticoids protect thymocytes from apoptosis⁶⁹; such antiapoptosis is also found with neural and other cells.^129–133

The physiologic significance of these effects is uncertain. Glucocorticoid-induced apoptosis has been invoked to account for immunosuppression, which, at least for short-term effects (hours or days), is better explained by actions on cytokines as described below. Apoptosis might serve to eliminate toxic or otherwise dangerous activated lymphocytes.¹³⁴ Several plausible ideas have been proposed for glucocorticoid involvement in positive or negative thymic selection of the T cell repertoire.⁶⁹ Thymocyte apoptosis, in contrast to many antiinflammatory and immunosuppressive reactions of glucocorticoids, requires GRs that dimerize (i.e., it is mediated by transactivation of genes through palindromic GREs).³⁴

Effects via Cytokines and Other Mediators, and via Their Receptors

Many suppressive effects of glucocorticoids on immune and inflammatory reactions appear to be due to inhibition of production or activity of cytokines, chemokines, inflammatory agents, certain hormones and neurotransmitters, and other mediators that are released during responses to LPS and other forms of stress.^108,135,136 Although many of these results were originally obtained with cell cultures, most have been observed in intact organisms.²⁶

These mediators form communication networks for defense mechanisms that respond to stress-induced challenges to homeostasis: cytokines and chemokines respond to infection, inflammatory agents to tissue damage, neurotransmitters to “fight or flight” encounters, and so forth. By blocking communication, glucocorticoids limit the stress response, preventing it from overshooting and damaging the organism. Most mediators in excess can be toxic, even lethal. Glucocorticoids limit not only production of mediators but sometimes their effects, such as TNF-α toxicity and responses of lymphocytes to IL-2 and of eosinophils to IL-3, IL-5, interferon (IFN)-γ, and granulocyte-macrophage colony-stimulating factor (GM-CSF).¹³⁷

Chemokines or chemotactic cytokines are produced locally in tissues, thus influencing traffic and homing of leukocytes by binding to G protein–coupled cell-surface receptors. Their secretion, probably stimulated by such cytokines as IL-1 and TNF-α, is dramatically increased during inflammation, resulting in recruitment of leukocytes to the inflamed site. Significant glucocorticoid effects on cell traffic are due to inhibition of chemokine secretion and cell adhesion molecules.

Not all mediators are suppressed by glucocorticoids. Annexin 1 (lipocortin 1) is induced by glucocorticoids.¹³⁸ It is antiinflammatory in several systems and may mediate inhibitory effects of glucocorticoids on release of ACTH from the pituitary.¹³⁹ Macrophage migration inhibitory factor (MIF) represents a special case because it antagonizes glucocorticoid actions.¹⁴⁰ In vivo, glucocorticoids raise MIF levels in plasma and in thymus, spleen, and other cells, and MIF in turn counteracts glucocorticoid effects.^136,141 Some mediators, like IL-10, are stimulated under some conditions and are suppressed under others. In the acute phase response, which involves the proinflammatory cytokines IL-1, IL-6, and TNF-α, glucocorticoids both potentiate induction by cytokines of certain acute phase proteins and suppress production of the cytokines.¹⁴²

Several mediators that are suppressed by glucocorticoids have receptors that, paradoxically, are induced by glucocorticoids. As discussed below, glucocorticoid induction of mediator receptors is a possible mechanism for some permissive effects.

Molecular mechanisms by which glucocorticoids control mediator production vary.^108,143 IL-1 production is blocked at the levels of transcription, translation, and secretion. TNF-α and GM-CSF appear to be blocked through increased degradation of their mRNAs. IL-2, IL-3, and possibly IFN-γ are blocked at the transcriptional level. Some, like prostaglandins and nitric oxide, may be suppressed because induction of the enzyme that synthesizes them is inhibited. Underlying many of these effects may be GR protein-protein cross-talk with NF-κB, AP-1, and other transcription factors,¹⁴³ which is consistent with the fact that most antiinflammatory actions are unaffected in transgenic mice with GRs that cannot dimerize.³⁴ Mitogen-activated protein kinase (MAPK) phosphatase 1 (MKP-1) is involved in MIF-glucocorticoid cross-talk.¹⁴¹ Numerous other mechanisms are being investigated.

Not only do glucocorticoids influence cytokines, but glucocorticoid actions are regulated by such cytokines as MIF, which was already mentioned, and IL-1, IL-2, TNF-α, IL-10, and IL-11.